Microsporidia are ubiquitous parasites causing infections in insects, fish, and mammals. Recently microsporidia have been demonstrated to infect humans. These organisms cause ophthalmic and gastrointestinal infections, primarily in patients with AIDS. Several genera of human pathogens have been cultivated in cell culture. Presently there are no data regarding the ability of the human pathogens Encephalitozoon intestinalis and E. hellem to survive under various environmental conditions. Also, there are no reports regarding the effects of disinfectants on spores of these two species. The survival of microsporidial spores after exposure to disinfectants, such as chlorine, alcohol, and quaternary ammonium compounds, and environmental conditions, such as elevated temperature and dessication, will be studied. Cultivation of microsporidia in the shell vial system using various cell lines has been investigated and microsporidia appear to replicate in both fibroblast and epithelial cell lines. A monoclonal antibody produced by Dr. T. Nash (NIAID, LPD) has replaced Giemsa staining for detection of infected cells. Although the monoclonal antibody makes detection of infected cells easier, the assay is still cumbersome and tedious. In an effort to replace staining of infected cell cultures, a method for quantitation of microsporidial DNA has been developed. This assay uses polymerase chain reaction (PCR) and FRET probes in the Light Cycler Instrument to provide real time PCR data. This assay will provide us with a rapid and sensitive method to replace visual examination of infected cells using the monoclonal antibody.